Over the last four years, we have demonstrated that hyperhomocysteinemia (hHcys) contributes to the development of glomerular sclerosis associated with hypertension. The molecular mechanisms mediating the pathogenic action of hHcys can be dependent or independent on elevated arterial pressure. It has been shown that NADPH oxidase (NOX) activation independent of elevated arterial pressure initiates the cascade of podocyte injury and consequent glomerular sclerosis during hHcys. Now it is imperative to know how this cascade of glomerular injury is instigated and amplified by activated NOX and redox signaling. The goal of this grant proposal is to test whether the Nlrp3 inflammasome, an intracellular inflammatory machinery is a triggering mechanism for instigation and amplification of the cascade for glomerular injury in hHcys and to determine how NOX activation turns on inflammasomes and how activated inflammasomes work to cause podocyte injury and glomerular sclerosis. We hypothesize that Nlrp3 inflammasomes sense NOX-derived redox signals and thereby turn on the inflammatory response and damage podocytes in glomeruli, ultimately leading to glomerular sclerosis during hHcys. To test this hypothesis, three Specific Aims are proposed. Specific Aim 1 will determine whether activation of a redox sensor-Nlrp3 inflammasome contributes to podocyte injury and chronic glomerular inflammation and sclerosis in hHcys induced by the folate-free diet in vivo, using Nlrp3-/- mice with or without rescuing Nlrp3 gene and wild type mice with locally silenced Nlrp3 gene. Specific Aim 2 will address how Nlrp3 inflammasomes are activated in podocytes by increased Hcys levels with a focus on the roles of NOX-mediated redox signaling and corresponding mechanisms mediating its action. Specific Aim 3 will explore the mechanisms by which activation of Nlrp3 inflammasomes leads to podocyte injury and glomerular dysfunction or sclerosis by studying the actions of inflammasome products such as IL-1b, IL-18 and DAMPs in cultured podocytes and in mice with hHcys induced by the folate-free diet. To our knowledge, these proposed studies represent the first effort in the field to study the role of an intracellular inflammatory machiner, the Nlrp3 inflammasome, as a redox sensor to switch on glomerular inflammation and to directly induce podocyte injury during hHcys.